Dopamine (DA) is a key transmitter in motor and emotive pathways of the brain. Dysfunction of dopaminergic neurotransmission underlies a variety of brain disorders that have a critical impact on society, including Parkinson's and Huntington's diseases, schizophrenia, and addiction. DA cell groups in the midbrain provide the primary source of DA to the CNS. Cells of the substantia nigra pars compacta (SNc) project rostrally to innervate the dorsal striatum (nigrostriatal DA system) whereas those of the adjacent ventral tegmental area (VTA) project to the nucleus accumbens and other limbic structures (mesolimbic DA system). A potentially unique characteristic of DA neurons is that they release DA from their dendrites in midbrain, as well as from distant axon terminals. Dendritic, as well as terminal release of DA is critical for DA-mediated behaviors, yet little is known about factors that regulate DA release In midbrain. Using carbon-fiber microelectrodes and fast-scan cyclic voltammetry to detect evoked DA release in real-time in brain slices, we have found significant differences in the Ca2+-dependence and Ca2+-channels required for dendritic vs. terminal release. Further, we have found distinct patterns of regulation by glutamate and GABA acting at ionotropic receptors in SNc, VTA and striatum. Significantly, we also discovered a novel, endogenous regulator of the nigrostriatal DA release: the reactive oxygen species (ROS), H202. In this continuation, we will investigate the distinct Ca2+ dependence of dendritic DA release, with emphasis on contributions from synaptic input to DA cells and release of Ca2+ from intracellular stores (Aim I). We will also elucidate the role of ROS as modulators of dendritic and terminal DA overflow (Aim ll-IV). Aim II will examine the involvement of H202 and other ROS in nigrostriatal vs. mesolimbic DA systems. Aim Ill will investigate sources of ROS, including terminals and cells adjacent to DA release sites. Aim IV will test whether H202 inhibits dendritic DA overflow by causing hyperpolarization of DA cells; effects on membrane properties will be indicated by whole cell recording.